This paper introduces a new bio-assessment tool, the mini-mobile environmental monitoring unit (MMU). The MMU is a portable, lightweight, energy-efficient, miniaturized laboratory that provides a low-flow system for on-site exposure of aquatic animals to local receiving waters in a protected, controllable environment. Prototypes of the MMU were tested twice in week-long studies conducted during the summers of 2008 and 2009, and in a 12-day study in 2010. In 2008, fathead minnows and polar organic chemical integrative samplers (POCIS) were deployed downstream from the Hastings, Nebraska wastewater treatment plant (WWTP), a waterway known to contain estrogenic contaminants in biologically active concentrations. In 2009, minnows and POCIS were deployed downstream, upstream and within the Grand Island, Nebraska WWTP, a site where the estrogenic contaminants had been detected, but were found at levels below those necessary to directly impact fish. In 2010, an advanced prototype was tested at the Sauk Center, Minnesota WWTP to compare its performance with that of traditional fish exposure methods including caged fish and static-renewal laboratory testing of effluent. Results from the prototype illustrate the capabilities of the MMU and offer an inexpensive monitoring tool to integrate the effects of pollutant sources with temporally varying composition and concentration.